GENETIC INSIGHTS INTO THE Β-CASEIN (CSN2) LOCUS IN HOLSTEIN-FRIESIANS: A STEP TOWARDS A2 MILK PRODUCTION

Authors

  • A. Bekyarova Department of Animal Husbandry, Faculty of Veterinary Medicine, Trakia University, Stara Zagora, Bulgaria Author

DOI:

https://doi.org/10.15547/tjs.2026.01.002

Keywords:

β-casein, CSN2 gene, Holstein-Frisian cattle, A2 milk

Abstract

Milk plays a major role as protein source in human diet. Among the casein fractions, β-casein, encoded by the CSN2 gene, is characterized by the presence of several polymorphic variants. The most predominant types in dairy cattle are A1 and A2. Phylogenetic analyses confirm that the A2 variant is the original, while A1 is a result of a mutation that can lead to the release of a biologically active peptide (BCM-7) during digestion. The present study aims to assess the allele and genotype frequencies of the CSN2 locus in 30 Holstein-Friesian cows bred at the Academic Technological Complex of Trakia University – Stara Zagora, and to analyze the genetic equilibrium of the population according to the Hardy–Weinberg principle. The analysis was performed with the BovineSNP50 v3 Bead Chip and established the presence of 3 genotypes - A1A1, A1A2 and A2A2. The results exhibit an allele frequency of the A1 allele = 0.4 and of the A2 allele = 0.6, with a χ² test confirming that the population is in Hardy–Weinberg equilibrium (χ² = 0.37, df = 1, p > 0.05). The witnessed high proportion of heterozygous individuals (A1A2) provides an opportunity for selection towards the A2A2 genotype. These results are fundamental for the development of breeding programs aiming the production of A2 β-casein milk, which has a growing consumer demand and potential health benefits.

References

Sanjulián, L.; Fernández-Rico, S.; González-Rodríguez, N.; Cepeda, A.; Miranda, J. M.; Fente, C.; et al. The Role of Dairy in Human Nutrition: Myths and Realities. Nutrients . Multidisciplinary Digital Publishing Institute (MDPI). February 1, 2025

Visioli, F.; Strata, A. Milk, dairy products, and their functional effects in humans: A narrative review of recent evidence. Advances in Nutrition. American Society for Nutrition, pp 131–143, 2014.

Kapaj, A.; Deci, E. World milk production and socio-economic factors effecting its consumption. In Dairy in Human Health and Disease across the Lifespan; Elsevier Inc.,; pp 107–115. 2017

Felis, A.; Pica-Ciamarra, U.; Reyes, E. Dairy’s Development and Socio-Economic Transformation: A Cross-Country Analysis. World, 6(3), 105. 2025

Koynarski, T.; Zapryanova, D.; Atanasoff, A.; Nikolov, G.; Hristova, D.; Secer, F. S. SEASONAL FLUCTUATIONS OF INNATE IMMUNITY AMONG THREE PHYLOGENETICALLY DIVERSE FISH. FRESENIUS ENVIRONMENTAL BULLETIN, cilt.27, sa.12, ss.8529-8535. 2018

Singh, U.; Deb, R.; Alyethodi, R. R.; Alex, R.; Kumar, S.; Chakraborty, S.; et al. Molecular markers and their applications in cattle genetic research: A review. Biomarkers and Genomic Medicine. Elsevier Inc., pp 49–58. 2014

Bozhilova-Sakova, M.; Viryanski, D.; Teneva, A.; Koynarski, T.; Todorovska, E.; Dimitrova, I. Genetic Diversity Analysis of the Rhodope Shorthorn Cattle Breed Based on SSR Markers; Vol. 29. 2023

Gaydarska, V. Dairy Cattle Breeding – Status and Challenges. Animal Sciences, LIII, 1–2/2016. 2016 (Bg)

Atamer, Z.; Post, A. E.; Schubert, T.; Holder, A.; Boom, R. M.; Hinrichs, J. Bovine β-casein: Isolation, properties and functionality. A review. International Dairy Journal. Elsevier Ltd, , pp 115–125. March 1, 2017

Walstra, P. On the Stability of Casein Micelles. Journal of Dairy Science, 73(8), 1965–1979. 1990

Aschaffenburg, R.; Drewry, J.: Occurrence of Different Beta-Lactoglobulins in Cow’s Milk. Nature volume 176, pages 218–219. 1955

Sun, Y.; Ding, Y.; Liu, B.; Guo, J.; Su, Y.; Yang, X.; et al. Recent advances in the bovine β-casein gene mutants on functional characteristics and nutritional health of dairy products: Status, challenges, and prospects. Food Chemistry. Elsevier Ltd. June 15, 2024

Liu, W.; Li, M.; Ren, H.; Liu, N.: Comparison of the Function and Conformation of Human β-Casein and Bovine β-Casein by Spectroscopic Study. Spectroscopy and Spectral Analysis. 34. 2014

Hamulka, J.; Górnicka, M.; Berthold-Pluta, A.; Kalinowski, A.; Habanova, M.; Madej, D. Effect of the Consumption of Milk with Beta-Casein A2A2, Milk with Beta-Casein A1A2 and a Plant-Based Drink on Metabolic Health in Adults: Protocol IMPA-CT Study. Nutrients, 17(24). 2025

Li, X.; Spencer, G. W. K.; Ong, L.; Gras, S. L. Beta casein proteins – A comparison between caprine and bovine milk. Trends in Food Science and Technology. Elsevier Ltd, pp 30–43. March 1, 2022

Khan, A.; Thakur, M. S.; Kumar, I.; Barwar, R.; Vishvakarma, S.; Vyas, J. Unveiling A1 and A2 Beta-Casein Gene Variants: Insights into Indian Cattle Breeds. Journal of Livestock Science, 16(3), 622–632. 2025

Sebastiani, C.; Arcangeli, C.; Ciullo, M.; Torricelli, M.; Cinti, G.; Fisichella, S.; et al. Frequencies evaluation of β-Casein gene polymorphisms in dairy cows reared in central Italy. Animals, 10(2). 2020

González-Rodríguez, N.; Vázquez-Liz, N.; Rodríguez-Sampedro, A.; Regal, P.; Fente, C.; Lamas, A. The Impact of A1- and A2 β-Casein on Health Outcomes: A Comprehensive Review of Evidence from Human Studies. Applied Sciences (Switzerland). Multidisciplinary Digital Publishing Institute (MDPI) July 1, 2025.

Escobar-Charry, M. A.; Quintanilla-Carvajal, M. X. An overview of bovine beta-casomorphin-7 (b-BCM7) and its potential impact on microbiota regulation and digestive health. Journal of Functional Foods. Elsevier Ltd. May 1, 2025.

Bolat, E.; Eker, F.; Yılmaz, S.; Karav, S.; Oz, E.; Brennan, C.; et al. BCM-7: Opioid-like Peptide with Potential Role in Disease Mechanisms. Molecules. Multidisciplinary Digital Publishing Institute (MDPI) May 1, 2024.

de Vitte, K.; Kerziene, S.; Klementavičiūtė, J.; de Vitte, M.; Mišeikienė, R.; Kudlinskienė, I.; et al. Relationship of β-casein genotypes (A1A1, A1A2 and A2A2) to the physicochemical composition and sensory characteristics of cows’ milk. Journal of Applied Animal Research, 50(1), 161–166. 2022

Szyda, J.; Liu, Z.; Reinhardt, F.; Reents, R. Estimation of quantitative trait loci parameters for milk production traits in German holstein dairy cattle population. Journal of Dairy Science, 88(1), 356–367. 2005

Lin, C. Y.; McAllister, A. J.; Ng-Kwai-Hang, K. F.; Hayes, J. F. Effects of Milk Protein Loci on First Lactation Production in Dairy Cattle. Journal of Dairy Science, 69(3), 704–712. 1986

Becerra-Quiroz, A. L.; Rivera-Jacinto, M. A.; Cueva-Rodríguez, M.; Tayca-Saldaña, A.; Alvarez-García, W.; Quilcate-Pairazamán, C.; et al. Frequency of A1 and A2 alleles of the β-casein gene in cattle in the Cajamarca region of Peru. Open Veterinary Journal, 15(11), 5971–5981. 2025

Çak, B.; Dede, S.; Yüksek, V.; Yılmaz, O.; Demirel, A. F. Genetic variability of alpha-casein, beta-casein, and kappa-casein genes in Holstein-Friesian, Simmental and Brown Swiss cattle. Journal of the Hellenic Veterinary Medical Society, 76(1), 8589–8596. 2025

Farrell, H. M.; Jimenez-Flores, R.; Bleck, G. T.; Brown, E. M.; Butler, J. E.; Creamer, L. K.; et al. Nomenclature of the proteins of cows’ milk - Sixth revision. Journal of Dairy Science, 87(6), 1641–1674. 2004

Ardicli, S.; Aldevir, Ö.; Aksu, E.; Kucuk, K.; Gümen, A. Associations of bovine beta-casein and kappa-casein genotypes with genomic merit in Holstein Friesian cattle. Archives Animal Breeding, 67(1), 61–71. 2024

Bisutti, V.; Pegolo, S.; Giannuzzi, D.; Mota, L. F. M.; Vanzin, A.; Toscano, A.; et al. The β-casein (CSN2) A2 allelic variant alters milk protein profile and slightly worsens coagulation properties in Holstein cows. Journal of Dairy Science, 105(5), 3794–3809. 2022

Yamada, A.; Sugimura, M.; Kuramoto, T. Genetic polymorphism of bovine beta-casein gene in Japanese dairy farm herds. Animal Science Journal, 92(1). 2021

Kamiński, S.; Ruść, A.; Cieślińska, A. A Note on Frequency of A1 and A2 Variants of Bovine Beta-Casein Locus in Polish Holstein Bulls *. 2006

Cartuche-Macas, L. F.; Navarrete-Mera, J. F.; Gutiérrez-Reinoso, M. A.; García-Herreros, M. Differential A1/A2 β-casein (CSN2) gene-derived allelic and genotypic frequencies across Ecuadorian exotic dairy cattle breeds. Frontiers in Veterinary Science, 12. 2025

Truswell, A. S. The A2 milk case: A critical review. European Journal of Clinical Nutrition. May, pp 623–631. 2005

Bisutti, V.; Pegolo, S.; Giannuzzi, D.; Mota, L. F. M.; Vanzin, A.; Toscano, A.; et al. The β-casein (CSN2) A2 allelic variant alters milk protein profile and slightly worsens coagulation properties in Holstein cows. Journal of Dairy Science, 105(5), 3794–3809. 2022

Sawicka-Zugaj, W.; Chabuz, W.; Barłowska, J.; Mucha, S.; Kasprzak-Filipek, K.; Nowosielska, A. Analysis of the Frequency of the A1 and A2 Alleles in the Beta-Casein Gene and the A, B and E Alleles in the Kappa-Casein Gene in Local Cattle Breeds: Polish Red and Polish White-Backed. International Journal of Molecular Sciences, 26(5). 2025

Cieślińska, A.; Fiedorowicz, E.; Zwierzchowski, G.; Kordulewska, N.; Jarmołowska, B.; Kostyra, E. Genetic polymorphism of β-Casein gene in Polish red cattle—preliminary study of A1 and A2 frequency in genetic conservation herd. Animals, 9(6). 2019

Rowan, T. N.; Schnabel, R. D.; Decker, J. E. Uncovering the architecture of selection in two Bos taurus cattle breeds. Evolutionary Applications, 17(2). 2024

Scott, B. A.; Haile-Mariam, M.; MacLeod, I. M.; Xiang, R.; Pryce, J. E. Evaluating the potential impact of selection for the A2 milk allele on inbreeding and performance in Australian Holstein cattle. Frontiers in Animal Science, 4. 2023

Sebastiani, C.; Arcangeli, C.; Ciullo, M.; Torricelli, M.; Cinti, G.; Fisichella, S.; et al. Frequencies evaluation of β-Casein gene polymorphisms in dairy cows reared in central Italy. Animals, 10(2). 2020

Ardicli, S.; Aldevir, O.; Aksu, E.; Gumen, A. The variation in the beta-casein genotypes and its effect on milk yield and genomic values in Holstein-Friesian cows. Animal Biotechnology, 34(8), 4116–4125. 2023

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Published

2026-03-12

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Original Contributions

How to Cite

GENETIC INSIGHTS INTO THE Β-CASEIN (CSN2) LOCUS IN HOLSTEIN-FRIESIANS: A STEP TOWARDS A2 MILK PRODUCTION. (2026). TRAKIA JOURNAL OF SCIENCES, 24(1), 6. https://doi.org/10.15547/tjs.2026.01.002

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